PROJECT SUMMARY/ABSTRACT for Project III Voltage-gated sodium channels (VGSCs) are responsible for the action potential in nerve and muscle. VGSCs are typically comprised of one -subunit and one or more -subunits. Mammalian neurons can have seven kinds (or isoforms) of -subunits and four isoforms of -subunits, which give rise to a large assortment of possible /-combinations or subtypes. Furthermore, a given neuron can have more than one subtype of VGSCs, where a given subtype may be localized to particular parts of the neuron. This diversity presents a serious challenge to the study of these channels and the development of drugs that target them. Drugs (such as local anesthetics) and toxins (such as tetrodotoxin, TTX) that block VGSCs have been mainstays in pain medication and basic neuroscience research. With notable exceptions, these small molecules do not discriminate readily among the various subtypes of VGSCs. For example, TTX is a very potent but indiscriminate blocker of most of the VGSC subtypes found in brain neurons. In contrast, conopeptides that target VGSCs can distinguish among neuronal subtypes. This proposal aims to exploit such conopeptides. There are four known families of conopeptides that target VGSCs, wherein members in each family have a characteristic structural framework and mechanism of action. It was recently discovered that members of two of these families could discriminate among VGSCs that had the same -subunit but different -subunits. This observation is unprecedented, and this proposal will: 1) examine the mechanism(s) underlying how -subunits influence conopeptide-susceptibility; 2) determine whether -subunits also affect the activity of members of the other two families of conopeptides, and if so, how; 3) develop conopeptides with sharper VGSC-subtype specificities. Finally, a new family of conopeptides, with an amino acid sequence unrelated to any previously characterized conopeptide, has been discovered that blocks most VGSCs provided that they do not have either a 2- or 4-subunit; the mechanism of action of this peptide will be investigated. This study will mainly entail electrophysiological experiments on VGSCs exogenously expressed in X. laevis oocytes and VGSCs endogenously expressed in neurons of rodent dorsal root ganglia.